Despite intense interest in the generation of cellular and humoral immune responses against viral pathogens, the anatomy of the response is imperfectly understood as infection proceeds from peripheral tissues to the draining lymph node (LN). Inside the LN, differences in the cell types infected by particular viruses, or even in the location of virus-infected cells, can impact virus containment within the LN and the subsequent generation of the adaptive immune response. Once a response is achieved, the production and release of effector molecules in the tissue must be tightly balanced to achieve viral clearance rather than immune-mediated pathology. Historically, vaccine development has required little, if any, understanding of the precise mechanisms and microanatomy of antiviral immunity. Rationally designing effective vaccines for many remaining viruses will require detailed knowledge of the factors that control and eliminate infections. The goal of the Viral Immunity and Pathogenesis Unit is to make fundamental discoveries about the processes that shape the creation and execution of antiviral immunity. Examining the generation of antiviral immunity in the draining lymph node The LN is a highly organized structure containing both hematopoietic and stromal cells. Lymphatic endothelial cells line nodal sinuses, preventing the free passage of lymph-borne particulates from sinuses to the LN interior. While a series of conduits channel proteins into the center of the LN, only molecules smaller than 70 kDa can access the conduits, and transfer of these proteins out of the conduits is controlled by fibroblastic reticular cells. As much larger virions transported in lymph are first deposited into sinuses, subcapsular sinus (SCS) macrophages serve as an important line of defense against invading pathogens as they filter incoming infectious material. Any material that is not captured by cells within the LN exits through the efferent lymphatics to downstream LNs or eventually the blood. Thus, the draining LN plays an important role in limiting viral spread. Because of this filtering function, macrophages and dendritic cells (DCs) within or with access to LN sinuses can be infected by many viruses with high frequency. The accessibility of incoming virions by different immune cell subsets greatly influences both the priming of the adaptive immune response and the containment of virus within the LN. The Viral Immunity and Pathogenesis Unit is currently investigating how the anatomy of and cell types within the draining LN affect adaptive immunity and viral containment after infection with vaccinia (VACV) and Zika (ZIKV) virus. Understanding clearance and pathogenesis in virus-infected tissues Once activated after recognition of virally derived peptides in the draining LN, antiviral effector CD8+ T cells traffic to infected tissues and eliminate virus-infected cells using a number of mechanisms. After developing approaches utilizing multiphoton microscopy to image live, virus-infected mice, we have visualized CD8+ T cells entering and patrolling VACV-infected skin. Surprisingly, after epicutaneous VACV infection, anatomically distinct targets in virus-infected tissues are cleared by different immune effector cells. Although most infectious VACV is produced by large, immobile infected keratinocytes, CD8+ T cells are ineffective at killing keratinocytes and instead eliminate motile infected inflammatory monocytes located in the dermis surrounding infected keratinocytes. Extending these studies, we recently reported a mechanism used by CD8+ T cells to locate VACV-infected inflammatory monocytes in the skin. The CXCR3-ligand chemokines CXCL9 and CXCL10 are greatly upregulated in VACV-infected skin, and CXCR3 expression maximizes the ability of effector CD8+ T cells to kill VACV-infected cells. We are currently exploring other factors that influence T cell-mediated killing of virus-infected cells in the skin and mucosa. Additionally, the Viral Immunity and Pathogenesis Unit is investigating the role of skin-resident antigen presenting cells during viral dissemination and clearance, after both VACV and ZIKV infection. Together, these studies will provide an increased understanding of features shaping antiviral immunity in barrier tissues.